Neurodevelopment is an intricate and dynamic process involving gene-environment interactions, which result in a series of changes in gene expression, cellular function, circuit formation, neuronal morphology and behavior. Epigenetic and chromatin-based modifications mediate distinct cellular gene expression profiles in vivo without directly affecting the DNA sequence in response to environmental stimuli. Exciting new research indicates that in addition to nucleosomal remodeling and covalent histone modifications, eukaryotic cells generate variation in chromatin structure through the introduction of variant histone proteins. Emerging evidence indicates that H3.3 represents an ideal chromatin mark for efficiently profiling dynamic changes in the epigenomic landscape during critical periods of transcriptional maintenance and regulation. We therefore will explore potential novel links between H3.3 specific variant localization and subsequent downstream patterns of gene expression during longitudinal stages of neurodevelopment. Our central hypothesis is that -- environmental stimuli experienced during critical periods of neurodevelopment are associated with robust changes in tissue specific H3.3 deposition profiles, such that comprehensive longitudinal analyses of the H3.3 epigenome promise to provide fundamentally novel insight into the impact of early environmental influences on the development of altered vulnerabilities to mental disorders during adulthood. We seek to test this working hypothesis in the following three interrelated Specific Aims: 1) Identify longitudinal histone H3.3 localization patterns in brain throughout neurodevelopment. We aim to perform unbiased epigenome-wide (ChIP-seq) analyses of histone H3.3 variant localization in the developing and adult brain, with specific emphasis on brain regions located within the limbic circuitry (e.g. hippocampus and cortex), using tagged H3.3 knock-in mice or a specific H3.3 antibody under standard basal conditions. 2) Examine the impact of early life experiences on histone H3.3 regulation and relocalization in brain. We aim to examine the ability of early life environmental enrichment to redirect H3.3 deposition throughout the chromatin template, thereby influencing normal patterns of developmental gene regulation. 3) Determine the functional relevance of H3.3 deposition by means of regulated loss-of-function and gain-of-function studies. We aim to generate complementary mouse lines and tools to study, and conduct experiments with these animals to explore the biological role of H3.3 variant deposition in guiding developmentally influenced cognitive and social behaviors. Collectively our studies will provide novel and critical information regarding the epigenome across neurodevelopment. Identification of genes and gene networks with altered epigenetic status will help to better describe the "epigenetic landscape" during sensitive periods of brain development and pinpoint novel targets for therapeutic intervention related to mental disorders. PUBLIC HEALTH RELEVANCE: The overall goal of this multidisciplinary, collaborative proposal is to apply state-of the-art gene and chromatin arraying technologies to better understand epigenetic variations mediating normal neural development, as well as those acting during sensitive periods of neurodevelopmental that may predispose individuals to the onset of mental illness. Mental disorders are devastating illnesses, which impact a considerable number of Americans and for which there are few treatments that adequately reverse neuronal impairment. Epigenetic reprogramming via exchange of specific histone variants, especially during times in which cells in the brain are not replicating, may represent a central mechanism for regulation of gene expression during sensitive periods of brain development.